This proposal outlines an integrated training and research plan for Dr. Guo Huang to complete further academic training under the mentorship of Dr. Eric Olson and transition to an independent investigator specializing in the field of heart development and regeneration. The PI is currently a Life Sciences Research Foundation Fellow working on comparative studies of the cardiac injury response in mouse and zebrafish. The overall objective of the research proposal is to understand the regulatory mechanisms and functions of retinoic acid (RA) signaling in heart development and regeneration. Heart attacks are the leading cause of morbidity and mortality in industrialized countries. An interruption of blood flow and oxygen supply during a heart attack often leads to death and loss of heart muscle cells, scar formation and subsequent potentially life-threatening cardiac arrhythmias. We have very limited, if any, regeneration ability to regrow cardiac muscles, which is in great contrast with adult zebrafish and neonatal mice that can regenerate up to 15% of the heart. In both regeneration models, cardiomyocyte proliferation is believed to be the dominant mechanism. In both mammalian embryonic development and adult fish heart regeneration, the epicardium-derived retinoic acid (RA) and its downstream signaling pathways have been implicated to be essential in cardiomyocyte proliferation and regeneration. Intriguingly, although the RA synthesis pathway is reactivated in the adult mouse epicardium after injury, the downstream RA response post-cardiac injury seems to remain inactive. Understanding the regulation and function of RA signaling during development and after injury might provide us novel therapeutic targets for drug development to promote myocyte regeneration following heart attacks. In the research plan, aim 1 will delineate the transcriptional regulation of the rate-limiting enzyme RALDH2 for RA synthesis during embryonic heart development and post-ischemic injury responses. Aim 2 will define the function of Raldh2 in embryonic heart development and neonatal heart regeneration. Aim 3 will determine whether gain of RA responses in the adult mouse epicardium can promote heart regeneration after cardiac injury. Aim 4 will study zebrafish Raldh2 regulation and RA response in the epicardium during zebrafish heart regeneration. In the mentored phase, the aim 1 and aim 2 will be completed, and new mouse and zebrafish transgenic models will be generated for continued investigation towards aim 3 and aim 4 in the independent phase. The proposed work is closely relevant to NIH's mission in that the expected outcome will provide essential insights on the evolutionarily conserved pathways activated by cardiac injury and molecular components that are differentially regulated in the adult mammalian heart that may account for the loss of regeneration potential in human.